This research proposal is designed to investigate the mechanisms through which low serum Mg2+ contributes to the development of cardiovascular disease. Epidemiological studies have shown that decreased dietary intake of magnesium (Mg) is associated with increased risk for cardiovascular disease, and clinical studies show significant reductions in free ionized Mg2+ (not total Mg) in serum of patients hospitalized with cardiovascular problems. Low extracellular Mg2+ has been shown to increase vascular contractility. Our studies with vascular smooth muscle cell in vitro have shown that lowering extracellular free Mg2+ within the pathophysiological range also produces an increase in membrane lipid oxidation, in fatty acid saturation, and in the transcription factor, nuclear factor kappa B (NF-KB). The observed fatty acid changes appear due to oxygen free radical oxidation of double bonds of the fatty acids in the sn-2 (middle carbon of glycerol) position to form platelet- activating factor (PAF) like lipids. Ceramide, a sphingomyelin-derived second messenger, may be responsible for the increased NF-KB levels in these smooth muscle cells. This factor regulates genes involved in the inflammatory response, considered to be a central element of atherogenesis. These results indicate that Mg2+ protects against oxidative damage to membrane lipids. Vascular smooth muscle cells apparently respond to an atherogenic stimulus independently, and thus could amplify or modify signals from endothelial cells. We will measure in vitro changes in membrane lipid oxidation, in PAF-like lipids during atherogenesis and in oxygen free radical formation as a function of[Mg2+]0 in aortic segments and in cultured aortic smooth muscle and endothelial cells. PAF-like lipids will be isolated and identified and tested for their mitogenic activity on smooth muscle cells. We will test the hypothesis that an increase in intracellular ionized Ca2+ is responsible for increased reactive oxygen species as extracellular Mg2+ is lowered within the pathophysiological range. It is clear that low extracellular Mg2+ is a risk factor which contributes to cardiovascular pathology, and that an understanding of its mechanism of action should provide increased opportunities for arresting the complex process of atherogenesis.